Twelve major users request funds to establish a central integrated protein identification system which allows high-throughput analysis of protein spots from 2D gels. The instrumentation requested consists of a mass spectrometer, 2D gel system and associated accessories. This technology has been developed during the last five years in the laboratory of Dr. John R. Yates, III, at the University of Washington, Seattle. The experimentally determined, tandem mass spectrum (MS/MS) for a protein spot of interest is compared with the compute generated, theoretical tandem mass spectra for every possible peptide of the corresponding size present within existing sequence databases. This technique is particularly successful when used in an organism whose genome has been completely sequenced, or one for which a large EST database is available. It is also possible to identify regions of proteins that are conserved between species, or to perform direct sequencing from MS/MS spectra using the same instrumentation. Thus, mass- spectrometry will provide an extremely powerful and rapid method for determining the sequences of interesting proteins and identifying them as known or unknown, thereby forming the foundation for subsequent experiments to determine their function. The twelve investigators will make use of this instrumentation to pursue various biological questions which will enhance their NIH funded research. In addition this instrumentation will facilitate the research of many other investigators in the Health Sciences Center who require identification of only a few proteins. No comparable instrument is currently available at the Health Sciences Center. This instrumentation will be compliment by other new protein chemistry equipment purchased with funds from the Cancer Center, Dean's Office and a consortium of Departments. The group on this SGI has a history of collegial interactions, a background that argues well for the successful sharing of the instrumentation requested.